Genome-wide association studies have identified susceptibility loci for esophageal squamous cell carcinoma (ESCC). We conducted a meta-analysis of all single-nucleotide polymorphisms (SNPs) that showed nominally significant P-values in two previously published genome-wide scans that included a total of 2961 ESCC cases and 3400 controls. The meta-analysis revealed five SNPs at 2q33 with P< 5 × 10−8, and the strongest signal was rs13016963, with a combined odds ratio (95% confidence interval) of 1.29 (1.19–1.40) and P= 7.63 × 10−10. An imputation analysis of 4304 SNPs at 2q33 suggested a single association signal, and the strongest imputed SNP associations were similar to those from the genotyped SNPs. We conducted an ancestral recombination graph analysis with 53 SNPs to identify one or more haplotypes that harbor the variants directly responsible for the detected association signal. This showed that the five SNPs exist in a single haplotype along with 45 imputed SNPs in strong linkage disequilibrium, and the strongest candidate was rs10201587, one of the genotyped SNPs. Our meta-analysis found genome-wide significant SNPs at 2q33 that map to the CASP8/ALS2CR12/TRAK2 gene region. Variants in CASP8 have been extensively studied across a spectrum of cancers with mixed results. The locus we identified appears to be distinct from the widely studied rs3834129 and rs1045485 SNPs in CASP8. Future studies of esophageal and other cancers should focus on comprehensive sequencing of this 2q33 locus and functional analysis of rs13016963 and rs10201587 and other strongly correlated variants.
We report improved whole-genome shotgun sequences for the genomes of indica and japonica rice, both with multimegabase contiguity, or almost 1,000-fold improvement over the drafts of 2002. Tested against a nonredundant collection of 19,079 full-length cDNAs, 97.7% of the genes are aligned, without fragmentation, to the mapped super-scaffolds of one or the other genome. We introduce a gene identification procedure for plants that does not rely on similarity to known genes to remove erroneous predictions resulting from transposable elements. Using the available EST data to adjust for residual errors in the predictions, the estimated gene count is at least 38,000–40,000. Only 2%–3% of the genes are unique to any one subspecies, comparable to the amount of sequence that might still be missing. Despite this lack of variation in gene content, there is enormous variation in the intergenic regions. At least a quarter of the two sequences could not be aligned, and where they could be aligned, single nucleotide polymorphism (SNP) rates varied from as little as 3.0 SNP/kb in the coding regions to 27.6 SNP/kb in the transposable elements. A more inclusive new approach for analyzing duplication history is introduced here. It reveals an ancient whole-genome duplication, a recent segmental duplication on Chromosomes 11 and 12, and massive ongoing individual gene duplications. We find 18 distinct pairs of duplicated segments that cover 65.7% of the genome; 17 of these pairs date back to a common time before the divergence of the grasses. More important, ongoing individual gene duplications provide a never-ending source of raw material for gene genesis and are major contributors to the differences between members of the grass family.
Comparative genome sequencing of indica and japonica rice reveals that duplication of genes and genomic regions has played a major part in the evolution of grass genomes
Long-term noninvasive cell tracing by fluorescent probes is of great importance to life science and biomedical engineering. For example, understanding genesis, development, invasion and metastasis of cancerous cells and monitoring tissue regeneration after stem cell transplantation require continual tracing of the biological processes by cytocompatible fluorescent probes over a long period of time. In this work, we successfully developed organic far-red/near-infrared dots with aggregation-induced emission (AIE dots) and demonstrated their utilities as long-term cell trackers. The high emission efficiency, large absorptivity, excellent biocompatibility, and strong photobleaching resistance of the AIE dots functionalized by cell penetrating peptides derived from transactivator of transcription proteins ensured outstanding long-term noninvasive in vitro and in vivo cell tracing. The organic AIE dots outperform their counterparts of inorganic quantum dots, opening a new avenue in the development of fluorescent probes for following biological processes such as carcinogenesis.
Transcription activator-like effector nucleases (TALENs) are a powerful new approach for targeted gene disruption in various animal models, but little is known about their activities in Mus musculus, the widely used mammalian model organism. Here, we report that direct injection of in vitro transcribed messenger RNA of TALEN pairs into mouse zygotes induced somatic mutations, which were stably passed to the next generation through germ-line transmission. With one TALEN pair constructed for each of 10 target genes, mutant F0 mice for each gene were obtained with the mutation rate ranged from 13 to 67% and an average of ∼40% of total healthy newborns with no significant differences between C57BL/6 and FVB/N genetic background. One TALEN pair with single mismatch to their intended target sequence in each side failed to yield any mutation. Furthermore, highly efficient germ-line transmission was obtained, as all the F0 founders tested transmitted the mutations to F1 mice. In addition, we also observed that one bi-allele mutant founder of Lepr gene, encoding Leptin receptor, had similar diabetic phenotype as db/db mouse. Together, our results suggest that TALENs are an effective genetic tool for rapid gene disruption with high efficiency and heritability in mouse with distinct genetic background.
Cryptochromes are blue light receptors that mediate light regulation of gene expression in all major evolution lineages, but the molecular mechanism underlying cryptochrome signal transduction remains not fully understood [1, 2]. It has been reported that cryptochromes suppress activity of the multifunctional E3 ubiquitin ligase CONSTITUTIVE PHOTOMORPHOGENIC 1 (COP1) to regulate gene expression in response to blue light [3, 4]. But how plant cryptochromes mediate light suppression of COP1 activity remains unclear. We report here that Arabidopsis CRY2 (cryptochrome 2) undergoes blue light-dependent interaction with the COP1-interacting protein SUPPRESSOR OF PHYTOCHROME A 1 (SPA1) [5, 6]. We demonstrate that SPA1 acts genetically downstream from CRY2 to mediate blue light suppression of the COP1-dependent proteolysis of the flowering-time regulator CONSTANS (CO) [7, 8]. We further show that blue light-dependent CRY2-SPA1 interaction stimulates CRY2-COP1 interaction. These results reveal for the first time a wavelength-specific mechanism by which a cryptochrome photoreceptor mediates light regulation of protein degradation to modulate developmental timing in Arabidopsis.
Genetic factors are thought to play a role in development for colorectal carcinogenesis. ICAM-1 is a polymorphic gene, thus, the present study investigated the relationship between the polymorphisms of ICAM-1 and the susceptibility and phenotypical characteristics of colorectal cancer (CRC).
The polymorphisms at ICAM-1 exon 4 (G241R) and exon 6 (E469K) were detected by PCR with sequence-specific primers. The relationship between specific genotypes of ICAM-1 and differentiation of CRC was evaluated by the histological grade.
We showed only GG genotype of ICAM-1 individuals in either CRC or normal controls. The KK genotype of ICAM-1 K469E was found more frequently than in the controls (P < 0.05). Patients with well-differentiated CRC displayed the KK more frequently than those of poor differentiation (P < 0.05).
The findings indicate that polymorphisms of G241R are rare in Chinese population and that KK genotype of ICAM-1 K469E is significantly associated with well differentiation of CRC.
Escherichia coli strains causing postweaning diarrhea (PWD) and edema disease (ED) in pigs are limited to a number of serogroups, with O8, O45, O138, O139, O141, O147, O149, and O157 being the most commonly reported worldwide. In this study, a DNA microarray based on the O-antigen-specific genes of all 8 E. coli serogroups, as well as 11 genes encoding adhesion factors and exotoxins associated with PWD and ED, was developed for the identification of related serogroups and virulence gene patterns. The microarray method was tested against 186 E. coli and Shigella O-serogroup reference strains, 13 E. coli reference strains for virulence markers, 43 E. coli clinical isolates, and 12 strains of other bacterial species and shown to be highly specific with reproducible results. The detection sensitivity was 0.1 ng of genomic DNA or 103 CFU per 0.3 g of porcine feces in mock samples. Seventeen porcine feces samples from local hoggeries were examined using the microarray, and the result for one sample was verified by the conventional serotyping methods. This microarray can be readily used to screen for the presence of PWD- and ED-associated E. coli in porcine feces samples.
Salmonella is a major cause of food-borne disease in many countries. Serotype determination of Salmonella is important for disease assessment, infection control, and epidemiological surveillance. In this study, a microarray system that targets the O antigen-specific genes was developed for simultaneously detecting and identifying all 46 Salmonella O serogroups. Of these, 40 serogroups can be confidently identified, and the remaining 6, in three pairs (serogroups O67 and B, E1 and E4, and A and D1), need to be further distinguished from each other using PCR methods or conventional serotyping methods. The microarray was shown to be highly specific when evaluated against 293 Salmonella strains, 186 Shigella strains, representative Escherichia coli strains, and 10 strains of other bacterial species. The assay correctly identified 288 (98%) of the Salmonella strains. The detection sensitivity was determined to be 50 ng genomic DNA per sample. By testing simulated samples in a tomato background, 2 to 8 CFU per gram inoculated could be detected after enrichment. This newly developed microarray assay is the first molecular protocol that can be used for the comprehensive detection and identification of all 46 Salmonella O serogroups. Compared to the traditional serogrouping method, the microarray provides a reliable, high-throughput, and sensitive approach that can be used for rapid identification of multiple Salmonella O serogroups simultaneously.
The CAPRI and CASP prediction experiments have demonstrated the power of community wide tests of methodology in assessing the current state of the art and spurring progress in the very challenging areas of protein docking and structure prediction. We sought to bring the power of community wide experiments to bear on a very challenging protein design problem that provides a complementary but equally fundamental test of current understanding of protein-binding thermodynamics. We have generated a number of designed protein-protein interfaces with very favorable computed binding energies but which do not appear to be formed in experiments, suggesting there may be important physical chemistry missing in the energy calculations. 28 research groups took up the challenge of determining what is missing: we provided structures of 87 designed complexes and 120 naturally occurring complexes and asked participants to identify energetic contributions and/or structural features that distinguish between the two sets. The community found that electrostatics and solvation terms partially distinguish the designs from the natural complexes, largely due to the non-polar character of the designed interactions. Beyond this polarity difference, the community found that the designed binding surfaces were on average structurally less embedded in the designed monomers, suggesting that backbone conformational rigidity at the designed surface is important for realization of the designed function. These results can be used to improve computational design strategies, but there is still much to be learned; for example, one designed complex, which does form in experiments, was classified by all metrics as a non-binder.
Dihydromyricetin (DHM) is a major active ingredient of flavonoids compounds. It exhibited anticancer activity and induced apoptosis in human hepatocellular carcinoma HepG2 cells according to our previous data. In this study, we investigated whether p53 is involved in DHM-triggered viability inhibition and apoptosis induction in cancer cells. MTT [3-(4, 5-Dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide] assay was employed to evaluate the viability of HepG2 cells after DHM treatment. Meanwhile, p53 small interfering RNA (siRNA) was adopted to silence p53 expression. Protein level of p53 and Bax/Bcl-2 were evaluated by western blot analysis. Cell counting assay showed that DHM inhibited HepG2 cell growth effectively in a time- and dose-dependent manner. P53 expression was significantly increased after DHM treatment, whereas Bcl-2 was reduced potently. Furthermore, after co-treatment with Pifithrin-α (PFT-α, p53 inhibitor), Bcl-2 expression was reversed. The expression of Bax was no significant change, which was also observed after p53 silence. These findings defined and supported a novel function that DHM could induce human hepatocellular carcinoma HepG2 cells apoptosis by up-regulating Bax/Bcl-2 expression via p53 signal pathway.
Forkhead box transcription factor 1 (FOXM1) has been reported to overexpress and correlate with pathogenesis in a variety of human malignancies. However, little research has been done to investigate its clinical significance in gastric cancer.
We examined the expression of FOXM1 in 103 postoperational gastric cancer tissues and 5 gastric cell lines by immunohistochemistry and western blot analysis respectively. Data on clinic-pathological features and relevant prognostic factors in these patients were then analyzed. Moreover, the association of FOXM1 expression and chemosensitivity to docetaxel in gastric cancer cells was further explored.
Our study demonstrated that the level of FOXM1 expression was significantly higher in gastric cancer than in para-cancer tissues (P < 0.001) and normal gastric cell lines (P = 0.026). No significant association was found between FOXM1 expression and any clinical pathological features (P > 0.1). FOXM1 amplification was identified as an independent prognostic factor in gastric cancer (P = 0.001), and its affection is more significant in patients with tumor size larger than 5 cm (P = 0.004), pT3-4 (P = 0.003) or pIII-IV (P = 0.001). Additionally, shown to mediate docetaxel resistance in gastric cancers by our research, FOXM1 was revealed to alter microtubule dynamics in response to the treatment of docetaxel, and the drug resistance could be reversed with FOXM1 inhibitor thiostrepton treatment.
FOXM1 can be a useful marker for predicting patients’ prognosis and monitoring docetaxel response, and might be a new therapeutic target in docetaxel resistant gastric cancer.
FOXM1; Gastric cancer; Prognosis; Docetaxel resistance
Based on the previous research that oroxylin A can suppress inflammation, we investigated the hepatoprotective role of oroxylin A against CCl4-induced liver damage in mice and then studied the possible alteration of the activities of cytokine signaling participating in liver regeneration. Wild type (WT) mice were orally administrated with oroxylin A (60 mg/kg) for 4 days after CCl4 injection, the anti-inflammatory effects of oroxylin A were assessed directly by hepatic histology and indirectly by measuring serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) and Albumin. Proliferating cell nuclear antigen (PCNA) staining was performed to evaluate the role of oroxylin A in promoting hepatocyte proliferation. Serum IL-1β, TNF-α, IL-6 and IL-1Ra levels were measured by enzyme-linked immunosorbent assay (ELISA) and liver HGF, EGF, TNF-α, IL-6, IL-1Ra and IL-1β gene expression was determined by quantitative real-time PCR. The data indicated that the IL-6 and TNF-α mRNA of oroxylin A administered group significantly increased higher than the control within 12 hours after CCl4 treatment. Meanwhile, oroxylin A significantly enhanced the expression of IL-1Ra at the early phase, which indicated that oroxylin A could facilitate the initiating events in liver regeneration by increasing IL-1Ra which acts as an Acute-Phase Protein (APP). In addition, a lethal CCl4-induced acute liver failure model offers a survival benefit in oroxylin A treated WT mice. However, oroxylin A could not significantly improve the percent survival of IL-1RI−/− mice with a lethal CCl4-induced acute liver failure.
Our study confirmed that oroxylin A could strongly promote liver structural remodeling and functional recovery through IL-1Ra/IL-1RI signaling pathway. All these results support the possibility of oroxylin A being a therapeutic candidate for acute liver injury.
In order to search for new structural modification strategies on fluoroquinolones, we have designed and synthesized a series of fluoroquinolone derivatives by linking various hydrazine compounds to the C-3 carboxyl group of levofloxacin and assessed their anticancer activities. Several novel levofloxacin derivatives displayed potent cytotoxicity against the tested cancer cell lines in vitro. In the present study, we investigated the effect of 1-Cyclopropyl-6-fluoro-4-oxo-7- piperazin-1, 4-dihydro- quinoline- 3-carboxylic acid benzo [1,3] dioxol-5- ylmethylene- hydrazide (QNT11) on the apoptosis of human hepatocarcinoma cells in vitro.
The inhibition effects of QNT11 on cell proliferation were examined by MTT assay. Cell apoptosis was determined by TUNEL and DNA agarose gel electrophoresis method. The topoisomerase ΙΙ activity was measured by agarose gel electrophoresis using Plasmid pBR322 DNA as the substrate. Cell cycle progression was analyzed using flow cytometry in conjunction with ethanol fixation and propidium iodide staining. Mitochondrial membrane potential (△ψm) was measured by high content screening image system. The caspase-9, caspase-8, caspase-3, Bcl-2, Bax, CDK1, Cyclin B1and cytochrome c protein expressions were detected by Western blot analysis.
QNT11 showed selective cytotoxicity against Hep3B, SMMC-7721, MCF-7 and HCT-8 cell lines with IC50 values of 2.21 μM, 2.38 μM, 3.17 μM and 2.79 μM, respectively. In contrast, QNT11 had weak cytotoxicity against mouse bone marrow mesenchymal stem cells (BMSCs) with IC50 value of 7.46 μM. Treatment of Hep3B cells with different concentrations of QNT11 increased the percentage of the apoptosis cells significantly, and agarose gel electrophoresis revealed the ladder DNA bands typical of apoptotic cells, with a decrease in the mitochondrial membrane potential. Compared to the control group, QNT11 could influence the DNA topoisomerase IIactivity and inhibit the religation of DNA strands, thus keeping the DNA in fragments. There was a significant increase of cytochrome c in the cytosol after 24 h of treatment with QNT11 and a decrease in the mitochondrial compartment. Observed changes in cell cycle distribution by QNT11 treated might be caused by insufficient preparation for G2/M transition. In addition, QNT11 increased the protein expression of Bax, caspase-9, caspase-8, caspase-3, as well as the cleaved activated forms of caspase-9, caspase-8 and caspase-3 significantly, whereas the expression of Bcl-2 decreased.
Our results showed that QNT11 as a fluoroquinolone derivative exerted potent and selectively anticancer activity through the mechanism of eukaryotic topoisomerase II poisoning. The growth inhibition was in large part mediated via apoptosis-associated mitochondrial dysfunction and regulation of Bcl-2 signaling pathways.
Fluoroquinolone derivatives; Hepatocarcinoma cell line; DNA topoisomerase II; Mitochondrial dysfunction; Apoptosis
AIM: To explore the optimal steroid therapeutic strategy for autoimmune pancreatitis (AIP).
METHODS: This study was conducted retrospectively in two large institutions in China. Patients with clinically, radiologically and biochemically diagnosed AIP were enrolled. The performed radiological investigations and biochemical tests, the regimen of the given steroid treatment, remission and relapse whether with and without steroid therapy were analyzed.
RESULTS: Twenty-eight patients with AIP received steroid treatment, while 40 patients were treated surgically by pancreatoduodenectomy, distal pancreatectomy and choledochojejunostomy, radiofrequency ablation for the enlarged pancreatic head, percutaneous transhepatic biliary drainage and endoscopic biliary drainage. The starting oral prednisolone dose was 30 mg/d in 18 (64.3%) patients and 40 mg/d in 10 (35.7%) patients administered for 3 wk. The remission rate of AIP patients with steroid treatment (96.4%) was significantly higher than in those without steroid treatment (75%). Maintenance therapy (oral prednisolone dose 5 mg/d) was performed after remission for at least 6-12 mo to complete the treatment course. Similarly, the relapse rate was significantly lower in AIP patients with steroid treatment (28.6%) than in those without steroid treatment (42.5%). Steroid re-treatment was effective in all relapsed patients with or without steroid therapy.
CONCLUSION: Steroid therapy should be considered in all patients with active inflammatory phase of AIP. However, the optimal regimen still should be trailed in larger numbers of patients with AIP.
Autoimmune pancreatitis; Chinese population; Steroid therapy; Remission; Relapse
Branching is an important trait of plant development regulated by environmental signals. Phytochromes in Arabidopsis mediate branching in response to the changes in the red light:far-red light ratio (R:FR), the mechanisms of which are still elusive. Here it is shown that overexpression of CONSTANS-LIKE 7 (COL7) results in an abundant branching phenotype which could be efficiently suppressed by shade or a simulated shade environment (low R:FR). Moreover, col7 mutants develop shorter hypocotyls and COL7 overexpression lines develop longer hypocotyls in comparison with the wild type in low R:FR, indicating that COL7 acts as an enhancer of the shade avoidance response. In shade or transient low R:FR, transcriptional and post-transcriptional expression levels of COL7 are up-regulated and positively associated with rapid mRNA accumulation of PHYTOCHROME INTERACTING FACTOR 3-LIKE 1 (PIL1), a marker gene of shade avoidance syndrome (SAS). Taken together, the results suggest a dual role for COL7 which promotes branching in high R:FR conditions but enhances SAS in low R:FR conditions.
Branching; COL7; light signal transduction; phytochrome B; PIL1; shade avoidance response.
Activation of Sir2-orthologs is proposed to increase lifespan downstream of dietary restriction (DR). Here we describe an examination of the effect of 32 different lifespan-extending mutations and four methods of dietary restriction on replicative lifespan (RLS) in the short-lived sir2Δ yeast strain. In every case, deletion of SIR2 prevented RLS extension; however, RLS extension was restored when both SIR2 and FOB1 were deleted in several cases, demonstrating that SIR2 is not directly required for RLS extension. These findings indicate that suppression of the sir2Δ lifespan defect is a rare phenotype among longevity interventions and suggest that sir2Δ cells senesce rapidly by a mechanism distinct from that of wild-type cells. They also demonstrate that failure to observe life span extension in a short-lived background, such as cells or animals lacking sirtuins, should be interpreted with caution.
aging; replicative lifespan; longevity; yeast; epistasis
The blue light receptors cryptochromes mediate various light responses in plants. The photoexcited cryptochrome molecules undergo a number of biophysical and biochemical changes, including electron transfer, phosphorylation, and ubiquitination, resulting in conformational changes to propagate light signals. Two modes of cryptochrome signal transduction have been recently discovered, the CIB (cryptochrome-interacting basic-helix-loop-helix 1)-dependent CRY2 regulation of transcription and the SPA1/COP1 (SUPPRESSOR OF PHYA /CONSTITUTIVELY PHOTOMORPHOGENIC1)-dependent cryptochrome regulation of proteolysis. Both cryptochrome signaling pathways rely on blue light-dependent interactions between the cryptochrome photoreceptor and its signaling proteins to modulate gene expression changes in response to blue light, leading to altered developmental programs of plants.
Polycystic Kidney Disease (PKD) kidneys exhibit increased extracellular matrix (ECM) collagen expression and metalloproteinases (MMPs) activity. We investigated the role of these increases on cystic disease progression in PKD kidneys.
We examined the role of type I collagen (collagen I) and membrane bound type 1 MMP (MT1-MMP) on cyst development using both in vitro 3 dimensional (3D) collagen gel culture and in vivo PCK rat model of PKD.
We found that collagen concentration is critical in controlling the morphogenesis of MDCK cells cultured in 3D gels. MDCK cells did not form 3D structures at collagen I concentrations lower than 1 mg/ml but began forming tubules when the concentration reaches 1 mg/ml. Significantly, these cells began to form cyst when collagen I concentration reached to 1.2 mg/ml, and the ratios of cyst to tubule structures increased as the collagen I concentration increased. These cells exclusively formed cyst structures at a collagen I concentration of 1.8 mg/ml or higher. Overexpression of MT1-MMP in MDCK cells significantly induced cyst growth in 3D collagen gel culture. Conversely, inhibition of MMPs activity with doxycycline, a FDA approved pan-MMPs inhibitor, dramatically slowed cyst growth. More importantly, the treatment of PCK rats with doxycycline significantly decreased renal tubule cell proliferation and markedly inhibited the cystic disease progression.
Our data suggest that increased collagen expression and MMP activity in PKD kidneys may induce cyst formation and expansion. Our findings also suggest that MMPs may serve as a therapeutic target for the treatment of human PKD.
Collagen I; 3 dimensional (3D) collagen gel culture; Doxycycline; Matrix metalloproteinase; PCK rats; Polycystic kidney disease
To investigate the placental proteome differences between pregnant women complicated with gestational diabetes mellitus (GDM) and those with normal glucose tolerance (NGT).
We used two-dimensional electrophoresis (2DE) to separate and compare placental protein levels from GDM and NGT groups. Differentially expressed proteins between the two groups were identified by MALDI-TOF/TOF mass spectrometry and further confirmed by Western blotting. The mRNA levels of related proteins were measured by realtime RT-PCR. Immunohistochemistry (IHC) was performed to examine the cellular location of the proteins expressed in placenta villi.
Twenty-one protein spots were differentially expressed between GDM and NGT placenta villi in the tested samples, fifteen of which were successfully identified by mass spectrometry. The molecular functions of these differentially expressed proteins include blood coagulation, signal transduction, anti-apoptosis, ATP binding, phospholipid binding, calcium ion binding, platelet activation, and tryptophan-tRNA ligase activity. Both protein and mRNA levels of Annexin A2, Annexin A5 and 14-3-3 protein ζ/δ were up-regulated, while the expression of the Ras-related protein Rap1A was down-regulated in the GDM placenta group.
Placenta villi derived from GDM pregnant women exhibit significant proteome differences compared to those of NGT mothers. The identified differentially expressed proteins are mainly associated with the development of insulin resistance, transplacental transportation of glucose, hyperglucose-mediated coagulation and fibrinolysis disorders in the GDM placenta villi.
Resveratrol is a natural polyphenolic compound that has cardioprotective, anticancer and anti-inflammatory properties. We investigated the capacity of resveratrol to protect RAW 264.7 cells from inflammatory insults and explored mechanisms underlying inhibitory effects of resveratrol on RAW 264.7 cells.
Murine RAW 264.7 cells were treated with resveratrol (1, 5, and 10 µM) and/or LPS (5 µg/ml). Nitric oxide (NO) and prostaglandin E2 (PGE2) were measured by Griess reagent and ELISA. The mRNA and protein levels of proinflammatory proteins and cytokines were analysed by ELISA, RT-PCR and double immunofluorescence labeling, respectively. Phosphorylation levels of Akt, cyclic AMP-responsive element-binding protein (CREB), mitogen-activated protein kinases (MAPKs) cascades, AMP-activated protein kinase (AMPK) and expression of SIRT1(Silent information regulator T1) were measured by western blot. Wortmannin (1 µM), a specific phosphatidylinositol 3-kinase (PI3-K) inhibitor, was used to determine if PI3-K/Akt signaling pathway might be involved in resveratrol’s action on RAW 264.7 cells. Resveratrol significantly attenuated the LPS-induced expression of nitric oxide (NO), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in RAW 264.7 cells. Resveratrol increased Akt phosphorylation in a time-dependent manner. Wortmannin, a specific phosphatidylinositol 3-kinase (PI3-K) inhibitor, blocked the effects of resveratrol on LPS-induced RAW 264.7 cells activation. In addition, PI3-K inhibition partially abolished the inhibitory effect of resveratrol on the phosphorylation of cyclic AMP-responsive element-binding protein (CREB) and mitogen-activated protein kinases (MAPKs) cascades. Meanwhile, PI3-K is essential for resveratrol-mediated phosphorylation of AMPK and expression of SIRT1.
Conclusion and Implications
This investigation demonstrates that PI3-K/Akt activation is an important signaling in resveratrol-mediated activation of AMPK phosphorylation and SIRT1 expression, and inhibition of phosphorylation of CREB and MAPKs activation, proinflammatory mediators and cytokines production in response to LPS in RAW 264.7 cells.
This review focus on the phytochemical progress and biological studies of plants from the genus Balanophora (Balanophoraceae) over the past few decades, in which most plants growth in tropical and subtropical regions of Asia and Oceania, and nearly 20 species ranged in southwest China. These dioeciously parasitic plants are normally growing on the roots of the evergreen broadleaf trees, especially in the family of Leguminosae, Ericaceae, Urticaceae, and Fagaceae. The plants are mainly used for clearing away heat and toxic, neutralizing the effect of alcoholic drinks, and as a tonic for the treatment of hemorrhoids, stomachache and hemoptysis. And it has been used widely throughtout local area by Chinese people.
Cinnamic acid derivative tannins, possessing a phenylacrylic acid derivative (e. g. caffeoyl, coumaroyl, feruloyl or cinnamoyl), which connected to the C(1) position of a glucosyl unit by O-glycosidic bond, are the characteristic components in genus Balanophora. In addition, several galloyl, caffeoyl and hexahydroxydiphenoyl esters of dihydrochalcone glucosides are found in B. tobiracola, B. harlandii, and B. papuana. Other compounds like phenylpropanoids, flavonoids, terpenoids and sterols are also existed. And their biological activities, such as radical scavenging activities, HIV inhibiting effects, and hypoglycemic effects are highlighted in the review.
Decoy receptor 3 (DcR3), a member of the tumor necrosis factor receptor (TNFR) superfamily, is associated with anti-tumor immunity suppression. It is highly expressed in many tumors, and its expression can be regulated by the MAPK/MEK/ERK signaling pathway. The MAPK/MEK/ERK pathway has been reported to be a regulator in tumor occurrence, development and clonal expansion. External-signal regulated kinase (ERK) is a vital member of this pathway.
The expression of DcR3 and ERK1/2 in tumor tissues of gastric cancer patients was significantly higher than the non-cancerous group (P < 0.05). There was no statistical difference among tumor tissues from patients with different ages or gender, and even of different differentiation (P > 0.05). However, in patients with stage I gastric cancer, the DcR3 and ERK1/2 levels were significantly lower than patients with more advanced stages.
DcR3 and ERK1/2 play a vital role in the development of gastric cancer, and they may be new markers for indicating the efficiency of gastric cancer treatment in the future.
Cancer cell vaccine-based immunotherapy has received increasing interest in many clinical trials involving patients with breast cancer. Combining with appropriate adjuvants can enhance the weak immunogenic properties of tumor cell lysates (TCL). In this study, diphtheria toxin (DT) and two tandem repeats of mycobacterial heat shock protein 70 (mHSP70) fragment 407-426 (M2) were conjugated to TCL with glutaraldehyde, and the constructed cancer cell vaccine was named DT-TCL-M2. Subcutaneous injection of DT-TCL-M2 in mice effectively elicited tumor-specific polyclonal immune responses, including humoral and cellular immune responses. High levels of antibodies against TCL were detected in the serum of immunized mice with ELISA and verified with Western blot analyses. The splenocytes from immunized mice showed potent cytotoxicity on Ehrlich ascites carcinoma cells. Moreover, the protective antitumor immunity induced by DT-TCL-M2 inhibited tumor growth in a mouse breast tumor model. DT-TCL-M2 also attenuated tumor-induced angiogenesis and slowed tumor growth in a mouse intradermal tumor model. These findings demonstrate that TCL conjugated with appropriate adjuvants induced effective antitumor immunity in vivo. Improvements in potency could further make cancer cell vaccines a useful and safe method for preventing cancer recurrence after resection.
Cancer immunotherapy; tumor cell lysates; cancer cell vaccine; breast cancer; adjuvant epitope
Immunoliposomes (ILs) anchored with internalizing human antibodies capable of targeting all subtypes of mesothelioma can be useful for targeted imaging and therapy of this malignant disease. The objectives of this study were to evaluate both the in vitro and in vivo tumor targeted internalization of novel internalizing human single chain antibody (scFv) anchored ILs on both epithelioid (M28) and sarcomatoid (VAMT-1) subtypes of human mesothelioma. ILs were prepared by post-insertion of mesothelioma-targeting human scFv (M1) onto preformed liposomes and radiolabeled with 111In (111In-IL-M1), along with control non-targeted liposomes (111In-CL). Incubation of 111In-IL-M1 with M28, VAMT-1, and a control non-tumorigenic cell-line (BPH-1) at 37°C for 24 h revealed efficient binding and rapid internalization of ILs into both subtypes of tumor cells but not into the BPH-1 cells; internalization accounted for approximately 81-94% of total cell accumulation in mesothelioma cells compared to 37-55% in control cells. In tumor bearing mice intravenous (i.v.) injection of 111In-IL-M1 led to remarkable tumor accumulation: 4 % and 4.7% injected dose per gram (% ID/g) for M28 and VAMT-1 tumors, respectively, 48 h after injection. Furthermore, tumor uptake of 111In-IL-M1 in live xenograft animal models was verified by single photon emission computed tomography (SPECT/CT). In contrast, i.v. injection of 111In-CL in tumor-bearing mice revealed very low uptake in both subtypes of mesothelioma, 48 h after injection. In conclusion, M1 scFv-anchored ILs showed selective tumor targeting and rapid internalization into both epithelioid and sarcomatoid subtypes of human mesothelioma, demonstrating its potential as a promising vector for enhanced tumor drug targeting.
Immunoliposomes; scFv antibody; mesothelioma; SPECT/CT imaging; tumor targeting
The molecular network sustained by different types of interactions among proteins is widely manifested as the fundamental driving force of cellular operations. Many biological functions are determined by the crosstalk between proteins rather than by the characteristics of their individual components. Thus, the searches for protein partners in global networks are imperative when attempting to address the principles of biology.
We have developed a web-based tool “Sequence-based Protein Partners Search” (SPPS) to explore interacting partners of proteins, by searching over a large repertoire of proteins across many species. SPPS provides a database containing more than 60,000 protein sequences with annotations and a protein-partner search engine in two modes (Single Query and Multiple Query). Two interacting proteins of human FBXO6 protein have been found using the service in the study. In addition, users can refine potential protein partner hits by using annotations and possible interactive network in the SPPS web server.
SPPS provides a new type of tool to facilitate the identification of direct or indirect protein partners which may guide scientists on the investigation of new signaling pathways. The SPPS server is available to the public at http://mdl.shsmu.edu.cn/SPPS/.